Growth Factor Binding Protein Research Articles

Eldecalcitol (ED-71)-induced exosomal miR-6887-5p suppresses squamous cell carcinoma cell growth by targeting heparin-binding protein 17/fibroblast growth factor-binding protein-1 (HBp17/FGFBP-1).

Heparin-binding protein 17/fibroblast growth factor-binding protein-1 (HBp17/FGFBP-1) was purified from A431 cell-conditioned media based on its capacity to bind to fibroblast growth factor 1 and 2 (FGF-1 and FGF-2). HBp17/FGFBP-1 has been observed to induce the tumorigenic potential of epithelial cells and is highly expressed in oral cancer cell lines and tissues. HBp17/FGFBP-1 is also recognized as a pro-angiogenic molecule as a consequence of its interaction with FGF-2. We have previously reported that Eldecalcitol (ED-71), an analog of 1α,25(OH)2D3, downregulated the expression of HBp17/FGFBP-1 and inhibited the proliferation of squamous cell carcinoma (SCC) cells in vitro and in vivo through NF-κb inhibition. To explore the possibility of microRNA (miRNA) control of HBp17/FGFBP-1, we analyzed exosomal miRNAs from medium conditioned by A431 cells treated with ED-71. Microarray analysis revealed that 12 exosomal miRNAs were upregulated in ED-71-treated A431 cells. Among them, miR-6887-5p was identified to have a predicted mRNA target matching the 3' untranslated region (3'-UTR) of HBp17/FGFBP-1. The 3'-UTR of HBp17/FGFBP-1 was confirmed to be a direct target of miR-6887-5p in SCC/OSCC cells, as assessed with a luciferase reporter assay. Functional assessment revealed that overexpression of miR-6887-5p in SCC/OSCC cells inhibited cell proliferation and colony formation in vitro, and inhibited tumor growth in vivo compared with control. In conclusion, our present study supports a novel anti-cancer mechanism involving the regulation of HBp17/FGFBP-1 function by exosomal miR-6887-5p in SCC/OSCC cells, which has potential utility as a miRNA-based cancer therapy.

Transgenerational Inheritance of Betaine-Induced Epigenetic Alterations in Estrogen-Responsive IGF-2/IGFBP2 Genes in Rat Hippocampus.

Betaine serves as a methyl donor for DNA methylation. Here, the effects of betaine on hippocampal expression of neurogenesis genes and their DNA methylation status across three generations are investigated. Pregnant rats (F0) are fed control and betaine-supplemented diets throughout gestation and lactation. Female F1 and F2 offspring at weaning, together with the F0 dams, are used in the study. Hippocampal expression of aromatase, estrogen receptor α, and estrogen-related receptor β is downregulated in F1, together with the estrogen-responsive insulin-like growth factor 2/insulin-like growth factor binding protein 2 (IGF-2/IGFBP2) genes. However, all these genes are upregulated in F2, which follows the same pattern of F0. In agreement with changes in mRNA expression, the imprinting control region (ICR) of IGF-2 gene is hypomethylated in F1 but hypermethylated in F2 and F0. In contrast, the promoter DNA methylation status of all the affected genes is hypermethylated in F1 but hypomethylated in F2 and F0. Methyl transfer enzymes, such as betaine homocysteine methyltransferase and DNA methyltransferase 1, follow the same pattern of transgenerational inheritance. These results indicate that betaine exerts a transgenerational effect on hippocampal expression of estrogen-responsive genes in rat offspring, which is associated with corresponding alterations in DNA methylation on ICR of IGF-2 gene and the promoter of affected genes.

Reprogramming of human fibroblasts into osteoblasts by insulin-like growth factor-binding protein 7.

The induced pluripotent stem cell (iPSC) is a promising cell source for tissue regeneration. However, the therapeutic value of iPSC technology is limited due to the complexity of induction protocols and potential risks of teratoma formation. A trans‐differentiation approach employing natural factors may allow better control over reprogramming and improved safety. We report here a novel approach to drive trans‐differentiation of human fibroblasts into functional osteoblasts using insulin‐like growth factor binding protein 7 (IGFBP7). We initially determined that media conditioned by human osteoblasts can induce reprogramming of human fibroblasts to functional osteoblasts. Proteomic analysis identified IGFBP7 as being significantly elevated in media conditioned with osteoblasts compared with those with fibroblasts. Recombinant IGFBP7 induced a phenotypic switch from fibroblasts to osteoblasts. The switch was associated with senescence and dependent on autocrine IL‐6 signaling. Our study supports a novel strategy for regenerating bone by using IGFBP7 to trans‐differentiate fibroblasts to osteoblasts.

Kbg Syndrome: Report of Musculoskeletal Motor Affection and Review of the Literature

Abstract Introduction: KBG syndrome is a rare autosomal dominant disorder. There are about 100 cases published in the literature. The most consistent shared characteristics are macrodontia, distinctive craniofacial findings, musculoskeletal abnormalities and development delays. Mutations in the ANKRD11 gene were identified in most patients and is an important diagnostic tool. Materials and Methods: We report a 16 years old boy with any interesting family background. He was evaluated by multiple specialist for multiple problems like transmission hearing loss, inguinoscrotal hernia, micrognathia, low set ears, large incisors, ogival palate…On the musculoskeletal field he has hypertonia, small hands and fingers on drumstick and a hip dysplasia since breastfeeding which was treated with Pavlik cast with good results. On the lower extremities we found asymmetric legs, bilateral knee flex and equine tendency, similar findings that those we found on PCI patients. Results: With all the findings, we send our patient to the Regional Reference Unit for Advanced Diagnosis of Rare Paediatric Diseases of Castilla y Leon. They have studied the ANKRD11 gene, with heterozygous result. Conclusion: To our knowledge, this is the first case reported in the literature giving importance to the musculoskeletal affectation in the KBG syndrome. We think that some of the features and restrictions we find in this patient are similar that those we can see in PCI, and the GMFM could be a great tool to measure and guide the treatment of this patients. Keywords Gross motor; KBG; Rare disease Abbreviations GMFM: Gross Motor Function Measure; EEG: Electroencephalogram; ADHD: Attention Deficit/Hyperactivity Disorder; GH: Growth Hormone; NGS: Next Generation Sequencing; CK: Creatinine Kinase; PCI: Children Cerebral Palsy; IGF-1: Insulin Like Growth Factor 1; TNN: Tenascin N; aCGH: Microarray Based Comparative Genomic Hybridization; IGFBP-3: Insulin Like Growth Factor Binding Protein 3; ANKRD11: ankyrin Repeat Domain 11

Global transcriptome analysis reveals relevant effects at environmental concentrations of cypermethrin in honey bees (Apis mellifera)

Cypermethrin is a frequently used insecticide in agriculture and households but its chronic and molecular effects are poorly known are . Here we describe effects of sublethal cypermethrin exposure on the global transcriptome in the brain of honey bees determined by RNA-sequencing. Exposure for 48 h to 0.3 ng/bee cypermethrin (3 ng/mL sucrose solution) causes 38 differentially expressed genes (DEGs), of which 29 are up-regulated and 9 down-regulated. Exposure to 3 ng/bee causes differential expression of 265 DEGs (209 up-, 56 down-regulated). Among the 24 DEGs shared by both concentrations are genes encoding muscular structure, muscular processes and esterase B1. Functional analysis (GO term analysis) confirms the enrichment of muscular development, structure and function among the 89 and 35 significantly altered GO terms at the low and high concentration, respectively. Up-regulation of nine DEGs determined by RT-qPCR showed a good correlation with RNA-sequence data. Among them are genes including esterase B1, titin, twitchin, mucin-19, insulin like growth factor binding protein, golgin like protein and helix loop protein. Our study demonstrates for the first time molecular effects of cypermethrin at environmental concentrations, which include expressional induction of genes encoding muscular and cellular processes and metabolism enzymes. Further studies should demonstrate the physiological consequences in bees.

Altered Angiogenic Gene Expression Profiles in Porcine Von Willebrand Disease

Objectives: Von Willebrand Factor (VWF) is a main factor of coagulation, but also known for its influence on angiogenesis by modifying the expression of several genes. The mechanisms were assumed to base on shared genetic pathways and also on an altered storage capability of certain proteins within the VWF-dependent Weibel-Palade Bodies (WPBs). As women affected by Von Willebrand Disease (VWD), a disease caused by a deficiency of VWF, may develop miscarriage and other complications during pregnancy which highly relies on proper angiogenesis, we analyzed the effect of VWD on the expression of genes potentially influenced by a deficiency of VWF within uterus tissue. Methods: Uterus tissue samples were taken from each two non-pregnant VWD pigs suffering from VWD type 1 (T1, heterozygous) and type 3 (T3, homozygous), resp., during late estrus. Two healthy pigs served as controls (WT, wild type). The genes F8 (coagulation factor VIII), FGA (fibrinogen alpha chain), FGB (fibrinogen beta chain), FGG (fibrinogen gamma chain), PLG (plasminogen), CXCL8 (C-X-C motif chemokine ligand 8), CALR (calreticulin), CTGF (connective tissue growth factor), and IGFBP7 (insulin like growth factor binding protein 7) were chosen for analysis and specific primers were designed. The expression of these genes was analyzed by qRT-PCR and relatively quantified against the endothelial specific housekeeping genes PROCR and CD31 using the ΔΔCT method and the Wilcoxon-Mann-Whitney statistic test. The differences in gene expression were compared among the genotypes. Results: F8, FGA, FGG, PLG, CXCL8, CTGF, and IGFBP7 showed no significantly altered expression profiles. However, genes showing differential expression with significant (p ≤ 0.05) or suggestive (p ≤ 0.1) p-values among the three genotypes in uterus tissue were CALR and FGB. CALR expression was reduced to 36.7% and 33.3% within uterus tissue of T1 and T3 pigs, respectively, compared to WT (p ≤ 0.1). When combining the VWD pigs of both types and comparing them to WT, their expression was reduced to 35.0% reaching significance (p ≤ 0.05). The expression of FGB was increased to about five-fold in T1 pigs compared to the WT (p ≤ 0.1), while it was not significantly changed in T3 individuals. Discussion and Conclusions: CALR protein is stored in the VWF-dependent WPBs and shows physical interactions with VWF. As a reduction of VWF goes along with a reduction of WPBs, adequate CALR release might be diminished. Additionally, CALR is known to inhibit endothelial cell proliferation and to strongly affect angiogenesis. FGB shows strong physical interactions with CALR. It is part of the coagulation system and a homozygous dysfunction of this gene causes afibrinogenemia and consequently bleeding disorders. The observed higher expression of FGB in T1 uterus tissue compared to the WT might therefore reflect a higher production of fibrinogen. However, this needs to be confirmed in further studies. The observed reduced expression of CALR in uterus tissue of VWD-affected pigs might therefore contribute to the antiangiogenic alterations observed in VWD patients and contribute to the pregnancy related complications in VWD. Disclosures No relevant conflicts of interest to declare.

IGFBP2 Plays an Essential Role in Cognitive Development during Early Life.

Identifying the mechanisms underlying cognitive development in early life is a critical objective. The expression of insulin‐like growth factor binding protein 2 (IGFBP2) in the hippocampus increases during neonatal development and is associated with learning and memory, but a causal connection has not been established. Here, it is reported that neurons and astrocytes expressing IGFBP2 are distributed throughout the hippocampus. IGFBP2 enhances excitatory inputs onto CA1 pyramidal neurons, facilitating intrinsic excitability and spike transmission, and regulates plasticity at excitatory synapses in a cell‐type specific manner. It facilitates long‐term potentiation (LTP) by enhancing N‐methyl‐d‐aspartate (NMDA) receptor‐dependent excitatory postsynaptic current (EPSC), and enhances neurite proliferation and elongation. Knockout of igfbp2 reduces the numbers of pyramidal cells and interneurons, impairs LTP and cognitive performance, and reduces tonic excitation of pyramidal neurons that are all rescued by IGFBP2. The results provide insight into the requirement for IGFBP2 in cognition in early life.

Novel markers of human ovarian granulosa cell differentiation toward osteoblast lineage: A microarray approach.

Under physiological conditions, human ovarian granulosa cells (GCs), are responsible for a number of processes associated with folliculogenesis and oogenesis. The primary functions of GCs in the individual phases of follicle growth are: Hormone production in response to follicle stimulating hormone (FSH), induction of ovarian follicle atresia through specific molecular markers and production of nexus cellular connections for communication with the oocyte. In recent years, interest in obtaining stem cells from particular tissues, including the ovary, has increased. Special attention has been paid to the novel properties of GCs during long-term in vitro culture. It has been demonstrated that the usually recycled material in the form of follicular fluid can be a source of cells with stem-like properties. The study group consisted of patients enrolled in the in vitro fertilization procedure. Total RNA was isolated from GCs at 4 time points (after 1, 7, 15 and 30 days of culture) and was used for microarray expression analysis (Affymetrix® Human HgU 219 Array). The expression of 22,480 transcripts was examined. The selection of significantly altered genes was based on a P-value <0.05 and expression higher than two-fold. The leucine rich repeat containing 17, collagen type I α1 chain, bone morphogenetic protein 4, twist family bHLH transcription factor 1, insulin like growth factor binding protein 5, GLI family zinc finger 2 and collagen triple helix repeat containing genes exhibited the highest changes in expression. Reverse-transcription-quantitative PCR was performed to validate the results obtained in the analysis of expression microarrays. The direction of expression changes was validated in the majority of cases. The presented results indicated that GCs have the potential of cells that can differentiate towards osteoblasts in long-term in vitro culture conditions. Increased expression of genes associated with the osteogenesis process suggests a potential for uninduced change of GC properties towards the osteoblast phenotype. The present study, therefore, suggests that GCs may become an excellent starting material in obtaining stable osteoblast cultures. GCs differentiated towards osteoblasts may be used in regenerative and reconstructive medicine in the future.

Cytokine Production in Cholangiocarcinoma Cells in Response to Clonorchis sinensis Excretory-Secretory Products and Their Putative Protein Components.

Clonorchis sinensis is a carcinogenic human liver fluke that promotes hepatic inflammatory environments via direct contact or through their excretory-secretory products (ESPs), subsequently leading to cholangitis, periductal fibrosis, liver cirrhosis, and even cholangiocarcinoma (CCA). This study was conducted to examine the host inflammatory responses to C. sinensis ESPs and their putative protein components selected from C. sinensis expressed sequenced tag (EST) pool databases, including TGF-β receptor interacting protein 1(CsTRIP1), legumain (CsLeg), and growth factor binding protein 2 (CsGrb2). Treatment of CCA cells (HuCCT1) with the ESPs or bacterial recombinant C. sinensis proteins differentially promoted the secretion of proinflammatory cytokines (IL-1β, IL-6, and TNF-α) as well as anti-inflammatory cytokines (IL-10, TGF-β1, and TGF-β2) in a time-dependent manner. In particular, recombinant C. sinensis protein treatment resulted in increase (at maximum) of ~7-fold in TGF-β1, ~30-fold in TGF-β2, and ~3-fold in TNF-α compared with the increase produced by ESPs, indicating that CsTrip1, CsLeg, and CsGrb2 function as strong inducers for secretion of these cytokines in host cells. These results suggest that C. sinensis ESPs contribute to the immunopathological response in host cells, leading to clonorchiasis-associated hepatobiliary abnormalities of greater severity.

IGFBP-2 stimulates calcium/calmodulin-dependent protein kinase kinase 2 activation leading to AMP-activated protein kinase induction which is required for osteoblast differentiation.

Insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding proteins-2 (IGFBP-2) function coordinately to stimulate osteoblast differentiation. Induction of AMP-activated protein kinase (AMPK) is required for differentiation and is stimulated by these two factors. These studies were undertaken to determine how these two peptides lead to activation of AMPK. Enzymatic inhibitors and small interfering RNA were utilized to attenuate calcium/calmodulin-dependent protein kinase kinase 2 (CaMKK2) activity in osteoblasts, and both manipulations resulted in failure to activate AMPK, thereby resulting in inhibition of osteoblast differentiation. IGFBP-2 and IGF-I stimulated an increase in CaMKK2, and inhibition of IGFBP-2 binding its receptor resulted in failure to induce CaMKK2 and AMPK activation. Injection of a peptide that contained the IGFBP-2 receptor-binding domain into IGFBP-2-/- mice activated CaMKK2 and injection of a CaMKK2 inhibitor into normal mice inhibited both CamKK2 and AMPK activation in osteoblasts. We conclude that induction of CaMKK2 by IGFBP-2 and IGF-I in osteoblasts is an important signaling event that occurs early in differentiation and is responsible for activation of AMPK, which is required for optimal osteoblast differentiation.

PF661 MOLECULAR PROFILING REVEALS HIGHLY DEREGULATED INSULIN GROWTH FACTOR BINDING PROTEINS IN PATIENTS WITH MYELOFIBROSIS AND RELATED NEOPLASMS

Background:The Insulin Growth Factor (IGF) system has been extensively studied in polycythemia vera (PV) showing erythroid progenitors to be hypersensitive to IGF1, levels of the IGF1‐binding protein – IGFBP‐1 to be fourfold increased but normal levels of IGF1 and IGFBP3. Furthermore, the stimulatory effect of IGFBP1 on erythroid progenitors in vitro provided a connection to overproduction of red blood cells in PV through IGF1. Most lately, increased IGF‐1R expression of mononuclear cells was found in patients with Philadelphia‐negative myeloproliferative neoplasms (MPN). In addition, the JAK2V617F mutation has been shown to result in IGF1 hypersensitivity in Ba/F3 cells providing a molecular basis for significance of IGF1 signaling in MPNs.Dysregulation of the IGF system is important in cancer development. We herein for the first time demonstrate several genes of IGF system to be markedly deregulated in MPNsAims:To study whole blood gene expression profiling in patients with MPNs.Methods:Gene expression microarrays were applied to whole blood from control subjects (n = 21) and patients with ET (n = 19), PV (n = 41), and PMF (n = 9). Affymetrix HG‐U133 2.0 Plus microarrays were used to generate gene expression profiles of 54.675 probe sets (38.500 genes). Total RNA was purified from whole‐blood, amplified to biotin‐labeled aRNA and hybridized to microarray chips. The statistical software R was applied to perform initial data processing and statistical analysis of microarray data.Results:20,439, 25,307, and 17,417 probe sets were significantly differentially expressed between control subjects and patients with ET, PV and PMF, respectively (FDR&lt;0.05). Several of the insulin‐like growth factor (IGF) and insulin‐like growth factor binding protein (IGFBP) genes were significantly deregulated (Table 1). IGF1, IGF2, IGFALS, IGFBP2, and IGF2BP1 were significantly upregulated in all three disease entities of which IGFBP2 was progressively and significantly upregulated from ET over PV to PMF (FDR&lt;0.05).Summary/Conclusion:In the present study we found several IGF‐related genes to be significantly deregulated in MPNs. Of note, IGF‐1 was significantly upregulated in all three subgroups of patients (ET, PV and PMF) and IGFBP1 was significantly downregulated in ET and PV patients. These observations are not in line with previous findings of normal IGF‐1 and highly elevated IGFBP1 levels in patients with PV. IGF‐1 has been shown to stimulate cell growth and up‐and down‐regulate the expression of anti‐ and pro‐apoptotic molecules, respectively, thereby protecting cancer cells from apoptosis. Enhanced IGF‐I gene expression might contribute to the chronic inflammatory state in MPN through enhancement of IL‐6 activity. It is intriguing to consider if IGF‐1 might also promote angiogenesis in MPN by stimulating VEGF secretion by e.g. megakaryocytes and monocytes similar to stimulation of VEGF‐secretion by e.g. myeloma cells. In conclusion, we have shown a number of genes in the IGF‐1 system to be deregulated in MPN, which altogether might contribute in driving clonal myeloproliferation, decreased apoptosis, bone marrow angiogenesis, immune evasion and inflammation as well. Future studies on the interplay between the transcriptome, IGF‐related proteins and IGF‐R expression levels might deliver new insight into the role of IGF‐system in the pathogenesis of MPNs and ultimately give the impetus to studies on the efficacy of inhibitors of IGF1 or IGF1R, which prohibit development of MPN‐phenotype in JAK2V617F mice.image

Mo1385 – Igf-1 (INSULIN LIKE GROWTH FACTOR 1) and Igfbp-2 (Insulin Like Growth Factor Binding Protein 2) Serum Levels As Potential Biomarkers in Differentiation Between Chronic Pancreatitis (CP) and Pancreatic Adenocarcinoma (PDAC) in Reference to Pancreatic Diabetes

Mo1385 – Igf-1 (INSULIN LIKE GROWTH FACTOR 1) and Igfbp-2 (Insulin Like Growth Factor Binding Protein 2) Serum Levels As Potential Biomarkers in Differentiation Between Chronic Pancreatitis (CP) and Pancreatic Adenocarcinoma (PDAC) in Reference to Pancreatic Diabetes

Insulin treatment does not affect follicular development but alters granulosa cell gene expression in dairy cows

The use of strategies to stimulate follicular growth are important, especially for use in timed artificial insemination (TAI) protocols, aiming to increase dairy cow's fertility. The aim of this study was to investigate the effect of insulin on follicular growth, steroid production and expression of genes related to follicular development. For this, cows were submitted to a progesterone (P4) and estradiol (E2) based synchronization protocol. In study 1, eleven primiparous lactating Holstein cows, received a single s.c. application of 0.25 IU/kg human insulin or no treatment (control) on D8 of the protocol. Blood samples were collected, and the dominant follicle diameter was assessed daily via transrectal ultrasonography, from D8 to D12. In study 2, eight multiparous non-pregnant and non-lactating Jersey cows, received a single s.c. application of 0.25 IU/kg human insulin, whereas cows from the control group received a single s.c. injection (1 mL) of saline solution (NaCl 0.9%). Blood samples were collected, and the dominant follicle diameter was assessed daily via transrectal ultrasonography from D6 to D9 of the protocol. Sixteen hours after insulin injection, follicular aspiration was performed. In study 1, insulin treatment decreased systemic glucose levels, but did not affect follicular growth. In study 2, the glucose decrease induced by insulin treatment was accompanied by a tendency of decreased progesterone levels in follicular fluid, along with a decrease in steroidogenic acute regulatory protein (STAR) and insulin like growth factor binding protein 2 (IGFBP2) mRNA abundance in granulosa cells. In conclusion, insulin treatment does not increase follicle growth and estradiol secretion in dairy cows, but decreases IGFBP2 and tends to increase pappalysin (PAPPA) mRNA abundance in granulosa cells, suggesting a positive effect on follicle development.

The Effect of Serum Igf-1,Igfbp-3 And Erythrocyte Transfusıons on Development of Mıld Retınopathy of Prematurıty

Introduction: Most important factors in retinopathy of prematurity (ROP) are prematurity and oxygen toxicity although blood transfusions, insulin like growth factor-1 ( IGF-1), insulin like growth factor binding protein 3 (IGFBP-3) and vascular endothelial growth factor (VEGF) also have important roles. The objectives of this study were, to measure IGF-1 and IGFBP-3 levels in preterm newborns before and after blood transfusion and assess if the effect of transfusion in development of ROP is via these mediators, and to investigate whether IGF-1 and IGFBP-3 levels measured at 32 and 33 gestational age (GA) were different in preterm newborns with and without ROP. Material and Methods: Preterm newborns with gestational age ≤34 weeks were included and blood samples were obtained before and after red blood cell (RBC) transfusion. Results: Thirty newborns were included, 17 of whom had ROP ( stage 1: n=11, stage 2: n=5, stage 3: n=1). IGF-1 and IGFBP-3 levels did not change after RBC tranfusion. Excluding the patient with stage 3 ROP all ROP patients were referred as mild ROP. No difference was observed between IGF-1 and IGFBP-3 levels of the patients with and without mild ROP. Patients with mild ROP had significantly more number of transfusions. Conclusions: Erythrocyte transfusion increased the frequency of ROP, whereas IGFBP-3 and IGF-1 were not associated of ROP.

Stem Cell-Derived Neurons as Cellular Models of Sporadic Alzheimer’s Disease

Alzheimer's disease (AD) occurs as either an autosomal dominant inherited disease or sporadically. While familial mutant genes can be expressed in cells or in animal models to assess dysregulated functions, sporadic AD cannot be replicated in models given our lack of understanding of causality. Furthermore, the study of sporadic forms of AD is difficult given the inaccessibility of brain tissues in living individuals and the manifestation of symptoms years after the onset of disease. Here, the objective was to assess if induced pluripotent stem cell-derived neurons from well-ascertained sporadic AD individuals could represent potential cellular models to determine the underlying molecular mechanisms of disease. We used cryopreserved peripheral blood mononuclear cells from three well-ascertained sporadic AD and three non-cognitively impaired (NCI) individuals of the CIMA-Q cohort to obtain iPSC-derived neurons. Microtubule associated protein 2 was decreased in AD neurons, whereas expression of AD-associated amyloid precursor protein, tau, and amyloid-β peptide was similar in AD and NCI individuals. RNA sequencing identified several upregulated and downregulated mRNAs in AD relative to NCI neurons. Of these, complement Factor H (CFH), signal regulatory protein beta1 (SIRPB1), and insulin like growth factor binding protein 5 (IGFBP5) were previously associated with AD. In addition, several transcription factors not previously associated with AD, but involved in neuronal proliferation and differentiation were differentially expressed. The results identify novel avenues for the study of the underlying causes of sporadic AD and support the establishment of additional lines to identify mechanisms of disease in sporadic AD individuals.

PKNOX2 suppresses gastric cancer through the transcriptional activation of IGFBP5 and p53

Promoter methylation plays a vital role in tumorigenesis through transcriptional silencing of tumor suppressive genes. Using genome-wide methylation array, we first identified PBX/Knotted Homeobox 2 (PKNOX2) as a candidate tumor suppressor in gastric cancer. PKNOX2 mRNA expression is largely silenced in gastric cancer cell lines and primary gastric cancer via promoter methylation. Promoter methylation of PKNOX2 was associated with poor survival in gastric cancer patients. A series of in vitro and in vivo functional studies revealed that PKNOX2 functions as a tumor suppressor. Ectopic PKNOX2 expression inhibited cell proliferation in GC cell lines and suppressed growth of tumor xenografts in mice via induction of apoptosis and cell cycle arrest; and suppressed cell migration and invasion by blocking epithelial-to-mesenchymal transition. On the other hand, knockdown PKNOX2 in normal gastric epithelial cells triggered diverse malignant phenotypes. Mechanistically, PKNOX2 exerts its tumor suppressive effect by promoting the up-regulation of Insulin like Growth Factor Binding Protein 5 (IGFBP5) and TP53. PKNOX2 binds to the promoter regions of IGFBP5 and TP53 and transcriptionally activated their expression by chromatin immunoprecipitation (ChIP)-PCR assay. IGFBP5 knockdown partly abrogated tumor suppressive effect of PKNOX2, indicating that the function(s) of PKNOX2 are dependent on IGFBP5. IGFBP5 promoted PKNOX2-mediated up-regulation of p53. As a consequence, p53 transcription target genes were coordinately up-regulated in PKNOX2-expressing GC cells, leading to tumor suppression. In summary, our results identified PKNOX2 as a tumor suppressor in gastric cancer by activation of IGFBP5 and p53 signaling pathways. PKNOX2 promoter hypermethylation might be a biomarker for the poor survival of gastric cancer patients.

The Diagnostic Value of Serum IGFBP7 in Patients with Esophageal Squamous Cell Carcinoma.

Esophageal squamous cell cancer (ESCC) is one of the leading malignant cancer in the world and especially in China with high incidence and mortality. The exploration of novel serum biomarkers is required for early detection of ESCC. We investigated the diagnostic value of serum insulin like growth factor binding protein 7 (IGFBP7) in ESCC, evaluating its potential to improve the diagnosis of ESCC. The serum samples of 106 patients with ESCC and 107 normal controls were tested by enzyme-linked immunosorbent assay (ELISA). The levels of IGFBP7 in ESCC group were significantly higher than that in normal controls, compared by the Mann-Whitney U test (P<0.0001). Using receiver operating characteristic (ROC) curve, the diagnostic value of serum IGFBP7 was demonstrated. Versus normal group, the area under the ROC curve (AUC) of all ESCC was 0.794 (95%CI: 0.735-0.853) and early-stage ESCC was 0.725 (95%CI: 0.633-0.817). With optimized cutoff value of 2.993 ng/mL, IGFBP7 showed certain diagnostic value with specificity of 90.7%, sensitivities of 40.6% and 32.4% in ESCC and early-stage ESCC, respectively. Considering the correlation between clinical data and IGFBP7, no significant association was found (all P>0.05). Thus, we supposed that serum IGFBP7 might be a potential biomarker in the diagnosis of ESCC.

Lack of impact of iodinated contrast media on kidney cell-cycle arrest biomarkers in critically ill patients

BackgroundIodinated contrast media may contribute to acute kidney injury. However, several recent works suggest that this toxicity is minimal in the clinical setting. Recently, urinary G1 cell-cycle arrest proteins tissue inhibitor of metalloproteinase 2 (TIMP-2) and insulin like growth factor binding protein 7 (IGFBP-7) were identified as highly sensitive and specific biomarkers for early detection of kidney aggression. The impact of contrast administration on those biomarkers has not been specifically evaluated but could provide clues about the toxicity of contrast media. This study aimed at measuring changes in TIMP-2 and IGFBP-7 urinary concentrations before and after a contrast-enhanced computed tomography in critically ill patients.Methods77 patients were included in a prospective observational cohort study. Urinary [TIMP -2]·[IGFBP-7] was measured before, 6 and 24 h after contrast infusion. Urine output and serum creatinine were followed 3 days.ResultsMedian [TIMP-2]·[IGFBP-7] was 0.06 [interquartile range 0.04;0.26], 0.07 [0.03;0.34] and 0.10 [0.04;0.37] (ng/mL)2/1000 respectively before, 6 and 24 h after contrast infusion. Individual changes from baseline were − 0.01 [− 0.11;0.11] and 0.00 [− 0.10;0.09] (ng/ml)2/1000 at 6 and 24 h. These changes were not higher among the patients increasing their Kidney Disease Improving Global Outcome (KDIGO) classification within 3 days after contrast infusion (n = 14 [18%] based on creatinine criterion only, n = 42 [55%] based on creatinine and urine output).ConclusionsChanges in [TIMP-2]·[IGFBP-7] urinary concentration after contrast-enhanced computed tomography were insignificant, suggesting minimal kidney aggression by modern iodinated contrast media.

Low‐quality rice straw forage increases the permeability of mammary epithelial tight junctions in lactating dairy cows

It is known that milking frequency and plasma hormones play important roles in regulating mammary permeability. However, it is still not known whether nutritional factors can influence udder permeability. This study was conducted to investigate mammary epithelial tight-junction permeability in lactating dairy cows fed different forage-based diets. Twenty mid-lactating dairy cows were allocated to ten blocks based on their parity and milk yield and then randomly assigned into rice straw-based diet and alfalfa-based diet groups. Both diets contained 15% corn silage and 55% concentrate (dry matter basis). In terms of forage sources, rice straw-based diets (RS) contained 30% rice straw, whereas alfalfa-based diets (AH) contained 23% alfalfa hay plus 7% Chinese wild rye hay. The concentrations of Na+ , Na+ /K+ ratio, bovine serum albumin, and plasmin in the milk, the plasma lactose concentration, and the mRNA abundance of BCL2 associated agonist of cell death, phosphatase and tensin homolog, and insulin like growth factor binding protein 5 in the mammary gland were greater in RS-fed cows than in AH-fed animals. Mammary expression of proliferating cell nuclear antigen and occludin was lower in RS-fed cows compared with the AH-fed group. The expressions of growth hormone receptor, claudin-1, -3, -4, and ZO-1 were similar in the two diet groups. The cows fed RS showed higher mammary alveolar permeability, likely due to its effect on proliferation/apoptosis rates of mammary epithelial cells. © 2018 Society of Chemical Industry.

Identifying the key genes and microRNAs in colorectal cancer liver metastasis by bioinformatics analysis and invitro experiments.

Colorectal cancer (CRC) is one of the principal causes of cancer-associated mortality worldwide. The high incidence of liver metastasis is the leading risk factor of mortality in patients with CRC, and the mechanisms of CRC liver metastasis are poorly understood. In the present study, 7 datasets, including 3 gene expression profile datasets and 4 microRNA (miRNA) expression profile datasets were downloaded from the NCBI Gene Expression Omnibus (GEO) database to identify potential key genes and miRNAs, which may be candidate biomarkers for CRC liver metastasis. Differentially expressed (DE) genes (DEGs) and DE miRNAs of primary CRC tumor tissues and liver metastatic CRC tumor tissues were selected using the GEO2R tool. Gene Ontology and Kyoto Encyclopedia of Gene and Genome pathway enrichment analyses were conducted using the Database for Annotation, Visualization and Integrated Discovery online database. Furthermore, Cytoscape with cytoHubba and the Molecular Complex Detection (MCODE) plug-in were used to visualize a protein-protein interaction (PPI) network for these DEGs, and to screen hub genes and gene modules in the PPI network. In addition, the online databases, TargetScan, miRanda, PITA, miRWalk and miRDB, were used to identify the target genes of the DE miRNAs. In the present study, 141 DEGs (97 upregulated and 44 downregulated) and 3 DE miRNAs (2 upregulated and 1 downregulated) were screened from the 3 gene expression microarray datasets and 4 miRNA expression microarray datasets, respectively. In total, 10 hub genes with a high degree of connectivity were selected from the PPI network, including albumin (ALB), coagulation factor II (F2), thrombin, apolipoprotein H (APOH), serpin family C member 1 (SERPINC1), apolipoprotein A1 (APOA1), α-1-microglobulin/bikunin precursor (AMBP), apolipoprotein C3 (APOC3), plasminogen (PLG), α-2 HS glycoprotein (AHSG) and apolipoprotein B (APOB). The most important module was detected in the PPI network using the MCODE plug-in. A total of 20 DEGs were identified to be potential target genes of these DE miRNAs, and novel miRNA-DEGs regulatory axes were constructed. In vitro experiments were performed to demonstrate that miR-885 promoted CRC cell migration by, at least partially, decreasing the expression of von Willebrand factor (vWF) and insulin-like growth factor binding protein 5 (IGFBP5). In conclusion, by using integrated bioinformatics analysis and in vitro experiments, key candidate genes were identified and novel miRNA-mRNA regulatory axes in CRC liver metastasis were constructed, which may improve understanding of the molecular mechanisms underlying CRC liver metastasis.